The underlying mechanism responsible for the development of diet-induced insulin resistance in skeletal muscle remains unresolved. Decreased insulin sensitivity is a key factor in the etiology of type 2 diabetes and, as such, identifying the underlying mechanism of insulin resistance is critical to devising appropriate prevention and treatment strategies. Recent evidence indicates high dietary fat intake increases mitochondrial hydrogen peroxide production and emission in muscle, shifting the intracellular redox environment to a more oxidized state. Blocking the hydrogen peroxide emission through the use of mitochondrial targeted antioxidants prevents the shift in cellular redox environment and preserves insulin sensitivity, providing evidence the mitochondrial respiratory system senses and initiates a counterbalance response to cellular nutritional overload. The long-term objectives of this project are to define the underlying bioenergetics mechanisms regulating mitochondrial hydrogen peroxide production/emission, to determine the impact on and integration with cellular redox systems, and to decipher the mechanism by which redox signaling networks link to the control of insulin sensitivity. The specific goals of this project ae to determine if flux through -oxidation is a primary factor governing mitochondrial hydrogen peroxide emission, cellular redox state, and insulin sensitivity; to determine the mechanism(s) regulating hydrogen peroxide production/emission by the pyruvate dehydrogenase complex; and to determine the potential role of hydrogen peroxide induced redox regulation of phosphatase activity as a potential mediator of diet-induced insulin resistance. State-of-the-art mitochondrial function analyses as well as gain- and loss-of-function mouse models will be employed to address these goals. It is anticipated these studies will reveal new insights regarding the underlying mechanism by which metabolic imbalance leads to insulin resistance in skeletal muscle, providing the framework for devising appropriately targeted prevention and/or treatment strategies. PUBLIC HEALTH RELEVANCE: This project seeks to define the underlying mechanism for the decrease in insulin sensitivity that occurs in skeletal muscle in response to nutritional overload, the primary event in the etiology of type 2 diabetes. This is highly significant as understanding the fundamental cause of insulin resistance is essential to devising appropriate preventive and/or treatment measures to reduce the health and financial impact of the obesity and diabetes epidemics.